Nanotechnology Tackles Brain Cancer

Transcripts - Nanotechnology Tackles Brain Cancer

1.
Brain cancer can be counted among
the most deadly and intractable diseases.
Often diagnosed after a patient exhibits
symptoms such as nausea, dizziness,
uncharacteristic behavior changes, or
paralysis, the growing mass of a brain
tumor will continue to squeeze out normal
tissue and degrade the brain's function if
left untreated. But treatment is elusive.
Tumors may be embedded in regions of
the brain that are critical to orchestrating
the body's vital functions, while they shed
cells to invade other parts of the brain,
forming more tumors too small to detect
using conventional imaging techniques.
Brain cancer's location and ability to
spread quickly makes treatment with
surgery or radiation like fighting an enemy
hiding out among minefields and caves,
and explains why the term "brain cancer"
is all too often associated with the word
"inoperable."
Making treatment even more challenging,
there is a system of blood vessels and
protective cells in the brain — the blood-
brain barrier — that admits only essential
nutrients and oxygen, and keeps out every-
thing else, including about 95 percent of
all drugs. This natural barrier puts serious
limits on how much a patient can benefit
from traditional chemotherapy and new
cancer drugs.
In an ideal situation, we would have a
"smart" drug that could cross the blood-
brain barrier, zero in on the cancer cells,
mark their location clearly for surgery, or
destroy them with such precision that it
would leave surrounding, normal brain
cells intact. Until now, such a scenario
seemed so far-fetched. But using nan-
otechnology, NCI-supported researchers at
the University of Michigan, the University
of Washington, the University of Texas
M. D. Anderson Cancer Center, Virginia
Polytechnic Institute, and Virginia
Commonwealth University are creating
ultrasmall particles that can target and
destroy cancer cells in the brain, even
those in tumors too small to be removed
surgically.
Getting Into the Brain — "Ticket"
Required
Among the properties of nanoparticles that
make them ideal candidates for recognizing
and treating brain cancer, their ability to
deliver a wide variety of payloads across
the blood-brain barrier is perhaps the most
important. Understanding how some
nanoparticles achieve this special "permis-
sion" to enter the brain requires a closer
look at how the blood-brain barrier works.
The blood-brain barrier permits the
exchange of essential nutrients and gases
between the bloodstream and the brain,
while blocking larger entities such as
microbes, immune cells and most drugs
from entering. This barrier system is a
perfectly logical arrangement, since the
brain is the most sensitive and complex
organ in the human body and it would not
make sense for it to become the battle-
ground of infection and immune response.
This biological "demilitarization zone" is
enforced by an elaborate and dense net-
work of capillary vessels that feeds the
brain and removes waste products. Each
capillary vessel is bound by a single layer
of endothelial cells, connected by "tight
junctions," thereby making it very difficult
for most molecules to exit the capillaries
and permeate into the brain.
Outside of the central nervous system,
capillaries have fenestra (the latin for
"window"), which are the cracks between
the cells in the vessel wall. Both small and
large molecules and even cells can leave
the capillary and enter into the surround-
ing tissue.
Instead of "leaking" material, brain capillary
walls closely regulate the flow of material
using molecular pumps and receptors that
recognize and transport nutrients such as
glucose, nucleosides, and specific proteins
into the brain. In other words, substances
need to be pre-recognized to enter.
So what allows some nanoparticles to get
into the brain? Nanoparticles that success-
fully cross the barrier are often coated with
polyethylene glycol (PEG), polysorbate, or
other polymer or surfactant (a detergent-
like substance). The exact mechanism of
nanoparticle transport into the brain is not
fully understood, but it is thought to
depend on the particle's size, material
composition, and structure. In some cases,
it appears that a specialized coating of
polymer or surfactant allows nanoparticles
to mimic molecules that would normally be
transported into the brain. For example,
polysorbate-coated nanoparticles are
thought to mimic low-density lipoproteins
(LDL), allowing them to be transported
across the capillary wall and into the brain
by hitching a ride on the LDL receptor.1
In another example, nanoparticles were
"decorated" with opioid peptides, short
pieces of protein that act as natural
painkillers. The opioid peptides bind to
specific receptors on the capillary walls,
which help carry the nanoparticles into the
brain.2
In other cases, no special tricks are
needed: larger tumors can disrupt the
local vasculature, creating leaky vessels
through which nanoparticles and other
molecules can easily penetrate.
Once inside the brain, a nanoparticle can
deliver a wide variety of payloads to detect
and treat cancer.
Throwing PEBBLEs at Brain Cancer
When the team of Raoul Kopelman,
Ph.D., at the University of Michigan,
thought of a tool to diagnose and treat the
most virulent forms of brain cancer, they
thought of pebbles. That is, 20 to 200
nanometer diameter nanoparticles they
dubbed Probes Encapsulated by
Biologically Localized Embedding
(PEBBLEs).3
NCI Alliance for Nanotechnology in Cancer Monthly Feature December 20051
Nanotechnology Tackles
Brain Cancer
NCI Alliance for Nanotechnology in Cancer Monthly Feature December 2005

2.
Kopelman designed the PEBBLEs to carry
a variety of agents on their surface, each
with a unique function. Therein lies another
major potential advantage of using nano-
particles to treat cancer: multifunctionality.
One target molecule immobilized on the
surface could guide the PEBBLE to a
tumor. Another agent could be used to help
visualize the target using magnetic
resonance imaging (MRI), while a third
agent attached to the PEBBLE could
deliver a destructive dose of drug or toxin
to nearby cancer cells. All three functions
can be combined in a single tiny polymer
sphere to make a potent weapon against
cancer (See Figure 1).
Kopelman introduced the common MRI
contrast element — gadolinium — to the
PEBBLEs. When injected into the blood-
stream, the nanoparticles wend their way
through the bloodstream. But because
they can transverse the blood-brain
barrier, and because they have a
targeting agent attached, the PEBBLEs
accumulate in the brain tumor —
enabling a clear MRI image within just
a few hours.
The next functional step is a remark-
able feat of nano-engineering. Each
PEBBLE carries a photocatalyst.
When stimulated by a
light source through a
micrometer-sized fiber-
optic probe inserted into
the skull, the photocata-
lyst converts oxygen into
a so-called singlet state,
which effectively
"bleaches" and destroys
nearby cells. The
PEBBLEs are inert and
harmless until the light
is turned on. Used in
combination with MRI
imaging, one could now
kill cancer cells at will,
while tracking the effec-
tiveness of the treatment
with imaging.
The targeted treatments
using nanoparticles may
offer a number of
advantages over tradi-
tional chemotherapy. In
chemotherapy, the drugs
permeate cells through-
out the body to damage
their DNA and prevent rapid growth, and
are only moderately more toxic to cancer
cells over normal cells. That is why patients
suffer so many side effects of chemo-
therapy including nausea, hair loss, and
anemia. In contrast, PEBBLEs are highly
localized to the cancer target, and do very
little damage to surrounding healthy tissue.
PEBBLEs and other nanoparticle drugs
could also avoid another serious problem
occurring in traditional chemotherapy —
for some cancers, over 50 percent of
patients become non-responsive to
treatment due to the development of
multi-drug resistance (MDR). MDR occurs
when cancer cells mutate and begin to
pump the chemotherapy drugs back out
before they can destroy the cell. The
cancer becomes "immune" to the drug.
But PEBBLEs act on the outside of the
cell, and the toxic payload of oxygen that
they deliver acts quickly, without giving the
cancer much chance to survive and
develop resistance.
In rat models of a type of brain cancer
called 9L-gliosarcoma, PEBBLE-based
treatment can significantly increase
survival time. Patients with this particularly
aggressive form of cancer rarely live more
than four months after diagnosis without
treatment. For rats, the clock runs out in
about five days. When rats were treated
with PEBBLEs targeted to 9L-gliosarcoma
tumors, some were still thriving after two
months, and an MRI image of their brain
revealed that the tumors had been
eliminated.
Kopelman and collaborators (including
Martin Philbert, Ph.D., at the University of
Michigan School of Public Health), hope
ultimately to prove the utility and safety of
this approach to treating brain cancer in
humans.
The Illuminated Brain
Unfortunately, the most common form of
primary brain cancer, glioblastoma, is also
the most aggressive and lethal. Glioblas-
toma tumors emerge rapidly and spread
throughout the brain. About 20,000
Americans are diagnosed each year, and
more than half of those patients will die
within 18 months. Surgery is limited in its
effectiveness because it is difficult to
differentiate visually between cancerous and
normal brain tissue, and any cancer cells
left behind are likely to proliferate and form
new tumors. In order to improve the odds
of eliminating all the cancer during surgery
and avoid removing healthy brain tissue,
researchers have devised a number of
fluorophores, or "glowing" molecules that
mark the tumor boundaries for removal.
NCI Alliance for Nanotechnology in Cancer Monthly Feature December 20052
NCI Alliance for Nanotechnology in Cancer Monthly Feature December 2005
Courtesy: Raoul Kopelman, Ph.D., University of Michigan
Figure1: The power of PEBBLEs is in their multifunctionality. One
tiny polymer sphere can contain a targeting agent that guides the
particle to cancer cells, a protective coating (PEG) that might also
help it cross the blood-brain barrier, photodynamic molecules that
catalyze the conversion of oxygen to highly reactive oxygen sin-
glets, magnetically dense metals for MRI contrast imaging, and a
fluorescent "beacon" to visually pinpoint the nanoparticles location.
Reprinted with permission from ref. 4; Copyright 2005 American Chemical Society
Figure 2: Schematic diagram for synthesis of nanoparticle-chlorotoxin (NPC) and NPC-Cy5.5 conjugates.
NPC-Cy5.5 is able to bind to and fluorescently illuminate glioblastoma tumors.

3.
But the fluorescent probes are difficult to
locate and use within the brain during
surgery. A multidisciplinary team at the
University of Washington and the Fred
Hutchinson Cancer Research Center in
Seattle, wanted to see if surgical outcomes
could be improved by a single probe that
accurately marks the location of a tumor
in pre-operative MRI scans, while guiding
the surgeon to those same locations in the
exposed brain.
To materials scientist Miqin Zhang, Ph.D.,
radiologist Raymond Sze, M.D., and oncol-
ogist Jim Olson, M.D., nanotechnology was
the perfect solution for creating such a
multifunctional probe.4
Starting with a 10
nanometer diameter iron oxide core that
serves as an MRI contrast material, Zhang
and colleagues coated the nanoparticles
with polyethylene glycol and modified
them with a fluorescent molecule called
Cy5.5. Cy5.5 gives off light at near-infrared
wavelengths, which — unlike visible light
— can penetrate several centimeters
through brain tissue (See Figure 2).
In order to selectively light up glioma
tumors through imaging, a targeting agent
had to be attached. Zhang and colleagues
selected chlorotoxin, a peptide derived
from the venom of the giant Israeli
scorpion, which binds specifically to a
tumor surface marker found in the vast
majority of gliomas. At 15 nanometers, the
final particle size and composition gave it
the best chance for crossing the blood-
brain barrier, and homing in on its target.
Tested on cultured cells, the nanoparticles
performed remarkably well (See Figure 3).
Studies clearly demonstrated uptake by
glioma cells, but not healthy brain cells,
and the nanoprobes were readily
detectable by both MRI and near-infrared
fluorescence. More work remains to be
done in animal studies, but Zhang and
coworkers are driven by a vision that some
day a surgeon will be able to use MRI to
map out a surgical plan, and then have a
visual guide for that plan in real time,
while operating on the brain.
According to Chun Li, Ph.D., at the
University of Texas M. D. Anderson
Cancer Center, and Eastman Kodak,
near-infrared emitting nanoparticles could
be a valuable tool for outlining the margins
of a tumor, and helping the surgeon to
avoid cutting into healthy tissue. Better
cameras and detection methods will be
needed before these nanoparticles are used
in a real operation, but a photographic
technology expert such as Eastman Kodak
seems ideal to address these challenges.
Kodak's engineering and chemistry teams
are backed by a long history of expertise in
dye chemistry. The company is supplying
high-quality near-infrared emitting nano-
particles with well-controlled properties.
Li's multidisciplinary approach, pooling the
expertise of engineers, chemists, and
biologists, is a good example of the multi-
disciplinary research collaborations needed
to move nanotechnology research forward
in the area of cancer.
Buckyballs Pack Heat
In the 1970s, a new variety of carbon was
discovered that formed hollow nanometer-
scale cage-like spheres. They were called
buckminsterfullerenes or buckyballs, after
the architect who built domed structures
of a similar shape, and were hypothesized
to have great potential for their ability to
carry atoms and molecules with useful
properties. Then in 1999, Virginia Tech
chemists Harry Dorn, Ph.D. and Harry
Gibson, Ph.D. created the first buckyballs
encapsulating rare earth metals, the kind
that can easily be recognized as a contrast
agent for MRI. The attachment of target-
ing agents to the outside of the carbon
cage allowed the buckyballs to accumulate
in tumors.
Now, Panos Fatouros, Ph.D., and neuro-
surgeon William Broaddus, M.D., Ph.D.,
both at Virginia Commonwealth University,
are collaborating with Dorn and Gibson
on a project using buckyballs to improve
the ability of MRIs to locate brain tumors,
and deliver a payload of radiation to
destroy them. Experiments on rats have
shown that buckyballs packed with the
MRI contrast metal gadolinium can
increase the sensitivity of MRI detection
by at least 40-fold. This level of precision
is reaching a point at which cancer cells
that have spread beyond the margins of
the tumor may become visible. Stray cells,
left behind after surgery, are thought to be
responsible for tumor relapse. Finding and
removing these cells could improve a
patient's chance of survival.
Fatouros and colleagues have created a
modified version of the buckyballs with a
fluorescent metal atom called terbium. A
glowing buckyball could guide surgeons to
remove tumors with greater precision.
Addition of yet another metal, lutetium,
would deliver a lethal dose of radiation to
the cancer cells, including those missed by
the surgeon. The research is about three to
five years away from testing in humans,
but the possibilities seem remarkable.
Detect, Treat, Track
There is a growing consensus that brain
cancer is a problem in need of a radically
different solution, and that nanotechnology
fits the bill. Functionalized nanoparticles
could provide precision detection, targeted
treatment, and real-time tracking that
conventional technology lacks. For a
disease in which only 5 percent to 32
percent of patients are likely to survive
after five years, large hope is riding on the
potential success of "small" technology.
Reprinted with permission from ref. 4; Copyright 2005 American Chemical Society
Figure 3: Fluorescent microscope images showing 9L-gliosarcoma cells light up with
chorotoxin-linked nanoparticles (red dots in B) but not when exposed to nanoparticles
without the chlorotoxin targeting agent (A).
NCI Alliance for Nanotechnology in Cancer Monthly Feature December 20053
NCI Alliance for Nanotechnology in Cancer Monthly Feature December 2005